Modulation of miR-146b by N6-methyladenosine modification remodels tumor-associated macrophages and enhances anti-PD-1 therapy in colorectal cancer

Purpose MicroRNA-146b (miR-146b) alleviates experimental colitis in mice by mediating macrophage polarization and the release of inflammatory factors. Our goals were to evaluate the antitumor efficacy of miR-146b in colorectal cancer (CRC) and to investigate the underlying mechanisms. Methods We used murine models of CRC to evaluate whether miR-146b influenced the progression of tumors independent of tumor-associated macrophages (TAMs). RNA immunoprecipitation, N6-methyladenosine (m(6)A) RNA immunoprecipitation and in vitro pri-miRNA processing assays were conducted to examine whether m(6)A mediates the maturation of pri-miR-146b/ miR-146b. In a series of in vitro and in vivo experiments, we further defined the molecular mechanisms of methyltransferase-like 3 (METTL3)/miR-146b-mediated antitumor immunity and its efficacy in combination with anti-PD-1 immunotherapy. Results We found that miR-146b deletion supported tumor progression by increasing the number of alternatively activated (M2) TAMs. Mechanistically, the m(6)A-related "writer" protein METTL3 and "reader" protein HNRNPA2B1 controlled miR-146b maturation by regulating the m(6)A modification region of pri-miR-146b. Furthermore, miR-146b deletion promoted the polarization of M2-TAMs by enhancing phosphoinositide 3-kinase (PI3K)/AKT signaling, and this effect was mediated by the class IA PI3K catalytic subunit p110 beta, which reduced T cell infiltration, aggravated immunosuppression and ultimately promoted tumor progression. METTL3 knockdown or miR-146b deletion induced programmed death ligand 1 (PD-L1) production via the p110 beta/PI3K/AKT pathway in TAMs and consequently augmented the antitumor activity of anti-PD-1 immunotherapy. Conclusions The maturation of pri-miR-146b is m(6)A- dependent, and miR-146b deletion-mediated TAM differentiation promotes the development of CRC by activating the PI3K/AKT pathway, which induces upregulation of PD-L1 expression, inhibits T cell infiltration into the TME and enhances the antitumor activity of anti-PD-1 immunotherapy. The findings reveal that targeting miR-146b can serve as an adjuvant to anti-PD-1 immunotherapy.